Cryptococcus neoformans is a ubiquitous encapsulated pathogenic yeast and a major cause of morbidity and mortality among patients with AIDS and other immunocompromising conditions. The natural reservoir of C. neoformans is exogenous, and cryptococcosis is acquired by inhalation of the desiccated yeast cells or basidiospores, which leads to dissemination and meningoencephalitis. Both clinical and environmental isolates of C. neoformans vary extensively in the expression of many phenotypes, including properties known to affect virulence. To address the evolution of this important opportunistic pathogen, we propose to investigate several basic aspects of the pathobiology of C. neoformans by analyzing the population genetics of both clinical and natural populations. In the previous period of support, we developed novel DNA markers to investigate the epidemiological and ecological relationships between natural and clinical populations of C. neoformans in patients with AIDS. We elucidated patterns of genetic variation, recombination and migration among populations. This proposal will extend those findings to determine mechanisms of evolution of C. neoformans and the source of cryptococcal infection in humans. Experimental studies will apply our molecular markers to investigate virulence traits. The three aims will (1) characterize the genetic structure and distribution of subpopulations of C. neoformans of serotypes A, D, and AD hybrids, (2) evaluate the genotypes and phenotypes of strains of C. neoformans that reflect their virulence, and (3) determine the affects of chromosomal rearrangements on the genotypes and phenotypes of C. neoformans.